microbeПроблемы особо опасных инфекцийProblems of Particularly Dangerous Infections0370-10692658-719XRussian Research Anti-Plague Institute “Microbe”10.21055/0370-1069-2024-3-74-80microbe-2043Research ArticleОРИГИНАЛЬНЫЕ СТАТЬИORIGINAL ARTICLESОсобенности везикуляции у вирулентных и авирулентных ЛПС-дефектных штаммов Francisella tularensis различной подвидовой принадлежностиPeculiarities of Vesiculation in Virulent and Avirulent LPS-Defective Strains of Francisella tularensis of Various Subspecieshttps://orcid.org/0000-0002-7772-9276АроноваН. В.AronovaN. V.

Аронова Надежда Валентиновна

Российская Федерация, 344002, Ростов-на-Дону, ул. М. Горького, 117/40

117/40, M. Gor’kogo St., Rostov-on-Don, 344002, Russian Federation

info@tularemia.ruhttps://orcid.org/0000-0001-8287-4294ПавловичН. В.PavlovichN. V.

Российская Федерация, 344002, Ростов-на-Дону, ул. М. Горького, 117/40

117/40, M. Gor’kogo St., Rostov-on-Don, 344002, Russian Federation

plague@aaanet.ruhttps://orcid.org/0000-0001-7268-9298МелоянМ. Г.MeloyanM. G.

Российская Федерация, 344002, Ростов-на-Дону, ул. М. Горького, 117/40

117/40, M. Gor’kogo St., Rostov-on-Don, 344002, Russian Federation

plague@aaanet.ruhttps://orcid.org/0000-0002-4091-649XЦимбалистоваМ. В.TsimbalistovaM. V.

Российская Федерация, 344002, Ростов-на-Дону, ул. М. Горького, 117/40

117/40, M. Gor’kogo St., Rostov-on-Don, 344002, Russian Federation

plague@aaanet.ruhttps://orcid.org/0000-0003-0550-2221НосковА. К.NoskovA. K.

Российская Федерация, 344002, Ростов-на-Дону, ул. М. Горького, 117/40

117/40, M. Gor’kogo St., Rostov-on-Don, 344002, Russian Federation

plague@aaanet.ruФКУЗ «Ростовский-на-Дону научно-исследовательский противочумный институт»РоссияRostov-on-Don Research Anti-Plague InstituteRussian Federation202429092024037480Copyright © Аронова Н.В., Павлович Н.В., Мелоян М.Г., Цимбалистова М.В., Носков А.К., 20242024Аронова Н.В., Павлович Н.В., Мелоян М.Г., Цимбалистова М.В., Носков А.К.Aronova N.V., Pavlovich N.V., Meloyan M.G., Tsimbalistova M.V., Noskov A.K.Данная работа распространяется под лицензией Creative Commons Attribution 4.0.This work is licensed under a Creative Commons Attribution 4.0 License.https://journal.microbe.ru/jour/article/view/2043

Целью работы явилось сравнительное изучение внеклеточных везикул (OMVs) и их композиционного состава у вирулентных и авирулентных (ЛПС -дефектных) штаммов Fracisella tularensis различной подвидовой принадлежности. Материалы и методы. С помощью трансмиссионной электронной микроскопии изучен процесс везикуляции у бактерий F. tularensis. Полученные препараты OMVs охарактеризованы по композиционному составу иммунологическими методами: реакция нейтрализации антител (РНА т), иммунохроматографический анализ, дот- и иммуноблоттинг. Результаты и обсуждение. Установлено, что все природные вирулентные штаммы, обладающие S-типом липополисахарида (ЛПС ), способны к образованию двух форм везикул: округлых и тубулярных (tubes), специфичных для F. tularensis. У авирулентных ЛПС -дефектных штаммов образование OMVs не зарегистрировано. Выявлено, что везикулы штаммов различных подвидов имеют свои индивидуальные морфологические особенности. Тубулярные структуры среднеазиатского штамма отличались бóльшими размерами по сравнению с голарктическим штаммом. Возможно, именно это определило тот факт, что препараты везикул F. tularensis subsp. mediasiatica, полученные с использованием фильтров с диаметром пор 0,22 мкм были резко обеднены тубулярными формами. Предложен способ обеззараживания суспензий бактерий с помощью гентамицина, не влияющий на морфологию и антигенную активность везикул. При сравнительном изучении нескольких серий концентрированных препаратов OMVs из разных штаммов туляремийного микроба обнаружено, что все образцы обладали антигенной активностью в реакции преципитации по Оухтерлони, РНА т, ИХ -тесте, дот- и иммуноблоттинге. Исследуемые образцы содержали иммунодоминантный антиген – ЛПС и несколько мажорных антигенактивных белков. Выявлены отличия в композиционном составе иммунодоминантных белков везикул, полученных из штаммов разных подвидов туляремийного микроба.

The aim of the study was to compare outer membrane vesicles (OMVs) and their composition in virulent and avirulent (LPS-defective) strains of Fracisella tularensis of various subspecies. Materials and methods. The vesiculation process in F. tularensis bacteria was studied using transmission electron microscopy. OMVs preparations were obtained and characterized by their composition using immunological methods: antibody neutralization test, immunochromatographic analysis, dot and immunoblotting. Results and discussion. It has been found that all natural virulent strains with the S type lipopolysaccharide (LPS) are able to produce two forms of vesicles – spherical and tubular (“tubes”), specific for F. tularensis. OMVs formation has not been registered in avirulent LPS-defective strains. It is revealed that vesicles of strains of various subspecies show their own individual morphological features. The tubes of F. tularensis subsp. mediasiatica are larger in size as compared to those of the subsp. holarctica strain. This is probably the reason why vesicle preparations from F. tularensis subsp. mediasiatica obtained using filters with a diameter of pores being 0.22 μm contained a significantly reduced number of tubular forms. A method of disinfection of bacterial suspensions using gentamycine, which does not affect the morphology and antigenic activity of vesicles, is proposed. A comparative study of several series of concentrated OMVs preparations obtained from different strains has revealed that all samples had antigenic activity in the precipitation Ouchterlony test, antibody neutralization test, immunochromatografic assay, dot-blot and immunoblotting. Immunodominant antigen – LPS and several major antigenic proteins have been detected in the preparations. Differences in the composition of vesicle proteins in the strains of various F. tularensis subspecies have been identified.

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The authors declare that there are no conflicts of interest present.